Studies in this project address the basic phenotype of the engrafting hematopoietic stem cell. Initial studies have shown that a cocktail of cytokines including IL-11, IL-3, IL-6 and steel factor or steel factor alone induce a profound engraftment defect in either normal host mice (male/female BALB/c transplant model) or lethally irradiated hosts (Ly5.1/Ly5.2 congenic transplant model or male/female BALB/c model). We plan to continue a characterization of the development of this engraftment defect, utilizing competitive transplantation of either male versus female or ly5.1 verses ly5.2 cells into appropriate lethally irradiated hosts. We will compete cytokine treated cells verses normal explant cells in these studies. Utilizing these models we will then assess whether or not we can create conditions in vitro allowing for the maintenance and survival of nonproliferating engrafting stem cells and whether utilizing such conditions we can take cytokine treated proliferating stem cells in vitro and quiesce them and reverse their phenotype to that of an engraftable stem cell. We are also carrying out synchronization studies utilizing Rhodaminelow Hoechstlow purified cells growth in cytokines and then subjected to various manipulation s to arrest cells in different points of cycle such as isoleucine deprivation or aphidocholin exposure in order to obtain highly synchronized populations of cells for study. The goal here would be to take dormant cells, stimulate their progression through cycle probably with steel factor and evaluate the capacity to engraft while dormant, in early G1, in S phase and (as a major goal of the project) in a second G1. Success to integrate retroviruses into proliferating stem cells and then return these cells to engraftability by reversing the engraftment defect.